instrucfions): Key components of bacterial pathogenesis in mammals are also involved in nematode killing. S. aureus kills C. elegans and that the C. elegans model can be used to study host-pathogen interactions involving staphylococcal biofilm exopolysaccharide. Importantly, S. aureus virulence determinants involved in mammalian pathogenesis, including the quorum-sensing global virulence regulatory system agr, the global virulence regulator sarA, anti the alternative sigma factor sigma(B) are required for full pathogenicity in nematodes. Based on these observations, we are developing a C. elegans-Staphylococcus aureus assay that is performed using 384-well plate technology. This model holds promise to overcome some of the main obstacles in current antibiofic discovery and may allow the identification of compounds that cannot be identified by in vitro screens. Our objective is to extend this C. e/egar?s-based assay by developing automated, high-throughput, whole animal screens for compounds with efficacy against methicillin-resistant S. aureus. Identifled compounds will be evaluated by carrying out dose-response and time-course studies, determining in vitro MICs, determining toxicity, and by prioritizing the compounds for tesfing in mammals. We will also test compounds identified previously in a C. elegans-E. faecalis screen for activity against E. faecalis and E. faecium. These experiments will validate the assay systems and demonstrate that these whole-animal assays can provide new lead compounds in real fime. The C. e/egans-based assays are particulariy appealing in that they should allow concurrent evaluafion of toxicity and antimicrobial activity as well as study of bacterial cells that are in a non-planktonic form. Thus false leads can be eliminated from further considerafion rapidly, and new leads will be cleariy identified. In addition to the compounds that have direct antibacterial activity, the C. e/ega/is-based assays may help identify compounds that affect staphylococcal virulence factors or immuno-modulate evolufionarily conserved elements ofthe host response. RELEVANCE (See instructions):